Expression, purification, crystallization and preliminary X-ray diffraction analysis of a ribokinase from the thermohalophile Halothermothrix orenii
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Hofmann, Andreas
Patel, Bharat KC
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Abstract
A ribokinase gene (rbk) from the anaerobic halothermophilic bacterium Halothermothrix orenii was cloned and overexpressed in Escherichia coli. The recombinant protein (Ho-Rbk) was purified using immobilized metal-ion affinity chromatography and crystals were obtained using the sitting-drop method. Diffraction data were collected to a resolution of 3.1 A ࠵sing synchrotron radiation. The crystals belonged to the orthorhombic space group P212121, with unit-cell parameters a = 45.6, b = 61.1, c = 220.2, and contained two molecules per asymmetric unit. A molecular-replacement solution has been found and attempts are currently under way to build a model of the ribokinase. Efforts to improve crystal quality so that higher resolution data can be obtained are also being considered.
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Acta Crystallographica Section F
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68
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Structural biology (incl. macromolecular modelling)